Monellin is an intensely sweet material present in the sap of "Serendipity Berries," the fruit of the West African plant, Dioscoreophyllum comminisii. The material has been purified to homogeneity and shown to be a basic protein with a molecular weight of about 1.1.times.10.sup.4 and is completely free of carbohydrate. Monellin is the first well characterized material among several sweet or taste modifying substances found in tropical plants. It has been characterized and shown to have two subunits of about the same size held together by non-covalent bonds. The two subunits are not identical and the flavor modifying ability of monellin is dependent upon the presence of both subunits and a single mercaptan group, which if blocked abolishes the sweetness.
Because of the uncertainties and cost of extracting natural products from plant sources, an alternative route to the production of protein sweeteners is of substantial interest. Recombinant techniques offer an opportunity to synthesize proteins of varying types. However, in employing recombinant techniques, one is required to develop a strategy for producing the gene, demonstrate successful expression of the protein in a cellular host, and isolate a product which is shown to have physiological activity. In many instances, it is necessary or desirable to modify the naturally occurring sequence, which substantially increases the uncertainties of success of the production of a useful product.
Relevant Literature
Morris et al., J. Biol. Chem. (1973) 248:534-439 describe the characterization of monellin. See also Cagan, Science (1973) 181:32-35; Wlodawer and Hodgson, Proc. Natl. Acad. Sci. USA (1975) 72:398-399; Bohak and Li, Biochimica et Biophysica Acta (1976) 427:153-170; Hudson and Bieman, Biochem. Biophys. Res. Comm. (1976) 71:212-220; Jirgenson, Biochim. Biophys. Acta (1976) 446:255-261; and Van der Wel and Loeve, FEBS Lett. (1973) 29:181-183 for further characterization. U.S. Pat. No. 3,998,798 describes the preparation of natural monellin.
Yeast leader sequences are described by Stack et al., Nucl. Acids Res. (1984) 12:6011-6030 and Julius et al., Cell (1984) 37:1075-1089. The GAL upstream activating sequence, a cis-acting DNA element, is described in Johnston and Davis, Mol. Cell. Biol. (1984) 4:1440-1448, while the transcription initiation region of glyceraldehyde-3-phosphate dehydrogenase is described by Holland and Holland, J. Biol. Chem. (1979) 254:5466-5475; 9839-9845; and Holland et al., ibid., (1983) 258:5291-5299.